A simple micro-growth assay for enumerating bacteria

A simple method for nonspecific determination of bacteria concentrations in a variety of liquid samples was developed. The assay was based on the time required for a sample grown in liquid media to reach a threshold turbidity. Samples were combined with media in a covered 96-well microwell plate and the turbidity was monitored in real time as the bacteria grew in a temperature-controlled plate reader. A significant problem with growth in microwells was condensation on the cover, which prevented accurate turbidity measurement. This problem was overcome by coating the cover with a small amount of surface-active agent. Salmonella and E. coli concentrations could be determined with a relative error of approximately 20% at levels from 10 to 10(6) cells/ml (eight replicates). An assay of 10 samples with standards required 10 min to set up and 20 min for data processing using a computer spreadsheet program. Growth time at 37 degrees C ranged from 4 h for samples at 10(7) cells/ml to 16 h for samples at 10 cells/ml.     

Identification of an alpha3-fucosyltransferase and a novel alpha2- fucosyltransferase activity in cercariae of the schistosome Trichobilharzia ocellata: biosynthesis of the Fucalpha1-->2Fucalpha1-- >3[Gal(NAc)beta1-->4]GlcNAc sequence

Fucose is a major constituent of the protein- and lipid-linked glycans of the various life-cycle stages of schistosomes. These fucosylated glycans are highly antigenic and seem to play a role in the pathology of schistosomiasis. In this article we describe the identification and characterization of two fucosyltransferases (FucTs) in cercariae of the avian schistosome Trichobilharzia ocellata, a GDP-Fuc:[Galbeta1-- >4]GlcNAcbeta-R alpha1-->3-FucT and a novel GDP-Fuc:Fucalpha-R alpha1-- >2-FucT. Triton X-100 extracts of cercariae were assayed for FucT activity using a variety of acceptor substrates. Type 1 chain (Galbeta1- ->3GlcNAc) based compounds were poor acceptors, whereas those based on a type 2 chain (Galbeta1-->4GlcNAc), whether alpha2'-fucosylated, alpha3'-sialylated, or unsubstituted, and whether present as oligosaccharide or contained in a glycopeptide or glycoprotein, all served as acceptor substrates. In this respect the schistosomal alpha3- FucT resembles human FucT V and VI rather than other known FucTs. N- ethylmaleimide, an inhibitor of several human FucTs, had no effect on the activity of the schistosomal alpha3-FucT, whereas GDP-beta-S was strongly inhibitory. Large scale incubations were carried out with Galbeta1-->4GlcNAc, GalNAcbeta1-->4GlcNAcbeta-O -(CH2)8COOCH3 and Fucalpha1-->3GlcNAcbeta1-->2Man as acceptor substrates and the products of the incubations were isolated using a sequence of chromatographic techniques. By methylation analysis and 2D-TOCSY and ROESY1H-NMR spectroscopy the products formed were shown to be Galbeta1-- >4[Fucalpha1-->2Fucalpha1-->3]GlcNAc, GalNAcbeta1-->4[Fucalpha1-- >2Fucalpha1-->3]GlcNAcbe ta-O-(CH2)8COOCH3, and Fucalpha1-->2Fucalpha1-- >3GlcNAcbeta1-->2Man, respectively. It is concluded that the alpha2- FucT and alpha3-FucT are involved in the biosynthesis of the (oligomeric) Lewisx sequences and the Fucalpha1-->2Fucalpha1-->3GlcNAc structural element that have been described on schistosomal glycoconjugates.      

Line Active Hybrid Lipids Determine Domain Size in Phase Separation of Saturated and Unsaturated Lipids

A simple model of the line activity of a hybrid lipid (e.g., POPC) with one fully saturated chain and one partially unsaturated chain demonstrates that these lipids preferentially pack at curved interfaces between phase-separated saturated and unsaturated domains. We predict that the domain sizes typically range from tens to hundreds of nm, depending on molecular interactions and parameters such as molecular volume and area per headgroup in the bulk fluid phase. The role of cholesterol is taken into account by an effective change in the headgroup areas and the domain sizes are predicted to increase with cholesterol concentration.     

Cytokinin Regulation of Flower and Pod Set in Soybeans (Glycine max(L.) Merr.)

Exogenous application of cytokinin to raceme tissues of soybean(Glycine max(L.) Merr.) has been shown to stimulate flower productionand to prevent flower abortion. The effects of these hormoneapplications have been ascertained for treated tissues, butthe effects of cytokinins on total seed yields in treated plantshave not been evaluated. Our objectives were to examine theeffects of systemic cytokinin applications on soybean yieldsusing an experimental line of soybeans, SD-87001, that has beenshown to be highly sensitive to exogenous cytokinin application.Soybeans were grown hydroponically or in pots in the greenhouse,and 6-benzylaminopurine (BA) was introduced into the xylem streamthrough a cotton wick for 2 weeks during anthesis. After theplants had matured, the number of pods, seeds per pod, and thetotal seed weight per plant were measured. In the greenhouse,application of 3.4 x 10^-7 moles of BA resulted in a 79% increasein seed yield compared with controls. Results of field trialsshowed much greater variability within treatments, with consistent,but non-significant increases in seed number and total yieldsof about 3%. Data suggest that cytokinin levels play a significantrole in determining total yield in soybeans, and that increasingcytokinin concentrations in certain environments may resultin increased total seed production. Copyright 2001 Annals ofBotany Company Glycine max, soybean, flower abortion, cytokinin, 6-benzylaminopurine, hydroponic, seed yield, wicking     

Cell lineage analysis of the Drosophila peripheral nervous system

The peripheral nervous system (PNS) of Drosophila provides a very well-characterized model system for studying the genes involved in basic processes of neurogenesis. Because of its simplicity and stereotyped pattern, each cell of the PNS can be individually identified and the phenotypic consequences of mutations can be studied in detail. Thus, some of the genetic mechanisms leading to the formation of type I sensory organs, the external, bristle-type sensory organs (es), and the internal, stretch-receptive chordotonal organs (ch) have been elucidated. Each sensory organ seems to be generated by a stereotyped pattern of cell division of individual ectodermal precursor cells. Recent advances in cell lineage analysis of the PNS have provided a detailed picture of almost all the lineages in the PNS, including those giving rise to the type II sensory neurons, also known as multiple dendritic (md) neurons. This knowledge will be instrumental in the precise characterization of the phenotypes associated with mutations in known and new genes and their interactions which determine cell fate decisions during neurogenesis. Here, we describe and compare three recently developed methods by which cell lineages have been assessed: single cell transplantation, bromodeoxyuridine (BrdU) incorporation studies, and the flp/FRT recombinase system from yeast. In the light of a more complete knowledge of the PNS lineages, we will discuss the effects of known mutations that alter neuronal cell fates. © 1996 Wiley-Liss, Inc.     

Interethnic genetic differentiation in Africa: HLA class I antigens in The Gambia.

A total of 752 individuals from The Gambia, west Africa who are representative of the major ethnic groups in the capital, Banjul, were serologically typed for HLA-A, -B, and -C antigens. Although all were typically "African" in their antigenic profiles, some marked frequency differences were found between the ethnic groups. Genetic distance comparisons with several other African populations showed that, although these west African populations clustered closely together, the positions of the various ethnic groups in The Gambia were consistent with historical and linguistic evidence of their affinities with one another and with other African populations. Despite the potential confounding effects both of selection by infectious diseases and of genetic drift caused by local differences in population structure, HLA frequencies appear to be of value in measuring inter- and intraregional population affinities in sub-Saharan Africa.